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rep-PCR-Mediated Genomic Fingerprinting: A Rapid and Effective Method to Identify Clavibacter michiganensis

First and eighth authors: MSU-DOE Plant Research Laboratory; first, eighth, and ninth authors: NSF Center for Microbial Ecology, Michigan State University, East Lansing 48824-1312; second, third, seventh, and ninth authors: Department of Botany and Plant Pathology, Michigan State University, Davis, CA 95616; fourth author: Department of Plant Pathology, University of California, Davis, CA 95616; fifth author: California Department of Food and Agriculture, Sacramento, CA 95814; and sixth author: Department of Bioagricultural Sciences and Pest Management, Colorado State University, Fort Collins 80523

The genomic DNA fingerprinting technique known as repetitive-sequence-based polymerase chain reaction (rep-PCR) was evaluated as a tool to differentiate subspecies of Clavibacter michiganensis, with special emphasis on C. michiganensis subsp. michiganensis, the pathogen responsible for bacterial canker of tomato. DNA primers (REP, ERIC, and BOX), corresponding to conserved repetitive element motifs in the genomes of diverse bacterial species, were used to generate genomic fingerprints of C. michiganensis subsp. michiganensis, C. michiganensis subsp. sepedonicus, C. michiganensis subsp. nebraskensis, C. michiganensis subsp. tessellarius, and C. michiganensis subsp. insidiosum. The rep-PCR-generated patterns of DNA fragments observed after agarose gel electrophoresis support the current division of C. michiganensis into five subspecies. In addition, the rep-PCR fingerprints identified at least four types (A, B, C, and D) within C. michiganensis subsp. michiganensis based on limited DNA polymorphisms; the ability to differentiate individual strains may be of potential use in studies on the epidemiology and host-pathogen interactions of this organism. In addition, we have recovered from diseased tomato plants a relatively large number of naturally occurring avirulent C. michiganensis subsp. michiganensis strains with rep-PCR fingerprints identical to those of virulent C. michiganensis subsp. michiganensis strains.

Additional keywords: bacterial canker of tomato , bacterial mosaic of wheat , ring rot of potato , taxonomy , wilt and leaf blight of corn , wilt of alfalfa .